Newly Synthesized S4MA Inhibits Inflammatory Responses Through STAT3/NF-κB Pathways in Human Colon Cancer HCT 116 Cells In Vitro

Background/Aim: This study aimed to investigate the impacts and underlying molecular mechanisms of E-4-(2-chloro-1-(4-methylbenzyl)-1H-indol-3-yl)but-3-en-2-one, C20H18ClNO (S4MA), a newly synthesized compound, on inflammation-associated cytokine responses in human colon cancer HCT 116 cells in vitro. Materials and Methods: Phase-contrast microscopy was utilized to detect the changes in cell morphology, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to analyze cell proliferation. Cytokine secretion was measured by enzyme-linked immunosorbent assay (ELISA), and mRNA levels were assessed by reverse transcriptase polymerase chain reaction (RT-PCR). Western blotting and confocal laser microscopy were employed to investigate the associated protein expression. Furthermore, the interaction of nuclear factor kappa-B (NF-κB) with specific DNA sequences in HCT 116 cells was evaluated using electrophoresis mobility shift assay (EMSA). Results: High concentrations of S4MA triggered cell morphological alterations in HCT 116 cells. Therefore, we reduced the concentration of S4MA to 1 μM for the inflammation-associated cytokine assay. S4MA at 0.5 and 1 μM inhibited cell proliferation. S4MA at 0.1 and 1 μM reduced the secretion of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) cytokines, and S4MA at 1 μM diminished the mRNA expression of IL-6 and TNF-α, as determined by ELISA and RT-PCR assays, respectively. Western blotting and confocal laser microscopy demonstrated that S4MA reduced the levels of signal transducer and activator of transcription 3 (STAT3)- and NF-κB-associated proteins in HCT 116 cells. Furthermore, S4MA reduced the interaction between NF-κB and its target DNA sequences within the nucleus, as determined by EMSA. Conclusion: S4MA inhibited inflammation-associated responses by regulating the STAT3/NF-κB pathways in human colon cancer HCT 116 cells. Therefore, S4MA may be a promising agent for suppressing inflammatory responses in future therapies for human colon cancer.